This invention relates to a method and apparatus for concentrating motile sperm from a sperm sample, including concentrating sperm in the vicinity of one or more oocytes for in vitro fertilization.
Infertility in man and animals can result from many causes and is frequently associated with low sperm counts, low sperm motility and low percentages of viable, motile sperm. One approach to overcoming these problems is in vitro fertilization ("IVF"), i.e., the combination of sperm and oocyte in a controlled and observable environment outside the body.
As presently practiced, in vitro fertilization generally involves the preliminary separation of healthy, motile sperm from less healthy or non-motile sperm by methods such as Percoll gradients or "swim-ups (Laufer et al., in Infertility, Serbel, M. M. eds., Appleton & Lange 1990)." Such fractionation steps are carried out in all cases, including those where sperm are normal (e.g., infertility due to pathology in the female's reproductive system). This measure is taken because even normal semen samples contain substantial numbers of immotile or dead sperm and epithelial cells, and these extraneous cells can interfere with interaction of the motile sperm with the eggs. In addition to removal of unwanted sperm, however these sperm fractionation techniques result in substantial losses of motile sperm as well, and thus introduce their own problems which may lead to the failure of in vitro fertilization. Indeed, in situations where sperm counts are very low (e.g. less than 10.sup.6 motile sperm per milliliter of ejaculate), fractionation may yield no detectable motile sperm.
Still another problem with standard sperm preparation techniques is that they all require centrifugation of the sample. Centrifugation can damage sperm (Aitken, R. J., and Clarkson, J. S., 1988. J. Androl. 9:367-376), and in some cases can lead to destruction of all competent sperm in a sample.
Once the sperm fraction to be used is obtained, the sperm are added to a sample dish containing a small number of oocytes (generally from 1 to 2) in liquid medium. Animal studies have shown that insemination in very small volumes of the liquid medium or "microdrops" can improve the efficiency of egg:sperm interaction and reduce the sperm:egg ratio needed for fertilization (Bavister, J. Exp. Zool. 210:259-264 1979). However, in human IVF, microdrops are not used because such drops are subject to rapid fluctuations of pH and/or changes in osmolarity due to evaporation. Moreover, the smaller the volume of a microdrop, the greater the requirement that competent sperm be separated from extraneous cells and debris. Thus, standard IVF procedures usually entail insemination of eggs in 2 milliliters of medium. Laufer et al.
These relatively large volumes necessitate insemination with large numbers of sperm (1-2.times.10.sup.5 or more) in order to assure fertilization. However, when the sperm sample is abnormal or when many eggs are obtained, the loss during fractionation can result in insufficient numbers of motile sperm for subsequent insemination. This problem is particularly severe when the sperm count is low and/or the percentage of motile sperm in the sample is low.
In vitro fertilization also includes methods whereby a sperm cell is directly injected into the egg. As with other IVF methods, fractionation (usually by Percall Gradients) is necessary in order to obtain individual motile sperm suitable for injection into the egg. Once again, where sperm counts are very low, fractionation may yield no sperm suitable for direct injection.
There thus exists a need in the art for a means for concentrating motile sperm from a sperm sample, for example for use in an in vitro fertilization technique which provides high fertilization efficiency without a preliminary sperm fractionation step being necessary. It is the object of the present invention to provide a method and apparatus to meet this need.